US 3868307 A
Distillers yeast is produced by a process which results in more rapid yeast growth, higher cell concentrations and greatly extended yeast viability. The process involves propagating yeast on a culture medium of cooked cereal grain mash which has been subjected to enzymatic action of glucamylase obtained from fungi of the genus Aspergillus. After propagation, as a result of substantially longer viability, the yeast can be stored in the culture medium for up to 3 days or longer with portions being withdrawn from time to time for inoculating distillery grain mashes.
Description (OCR text may contain errors)
United States Patent [1 1 Van Lanen, deceased et al.
[ Feb. 25, 1975 22 Filed:
[ PRODUCTION OF DISTILLERS YEAST  Inventors: James M. Van Lanen, deceased, late of Peoria, lll.; Merritt B. Smith; Weldon F. Maisch, both of Peoria, 111.
 Assignee: Hiram Walker & Sons, Inc., Peoria,
Aug. 15, 1973 [211 Appl. 110.; 388,359
Related US. Application Data  Continuation-impart of Ser. No.- 189,678, Oct. 15,
OTHER PUBLICATIONS Underkofler et al., Saccharification of Grain Mashes for Alcoholic Fermentation, Industrial and Engineering Chemistry, Vol. 38, 1946, (PP. 980-985) TPlA58.
Underkofler, et al., Industrial Fermentations, Vol. 1, Chem. Publ. Co., Inc., N.Y., 1954, (pp. 30-50) TP505u5.
Herstein et al., Chemistry and Technology of wines and Liquors, 2nd ed., D. Van Nostrand Co., Inc., New York, 1948 (pp. 75-78), TP505l-I48c.2.
Primary Examiner-David M. Naff Attorney, Agent, or Firml(irkland and Ellis  ABSTRACT Distillers yeast is produced by a process which results in more rapid yeast growth, higher cell concentrations and greatly extended yeast viability. The process involves propagating yeast on a culture medium of cooked cereal grain mash which has been subjected to enzymatic action of glucamylase obtained from fungi of the genus Aspergillus. After propagatiomas a result of substantially longer viability, the yeast can be stored in the culture medium for up to 3 days or longer with portions being withdrawn from time to time for inoculating distillery grain mashes.
6 Claims, 2 Drawing Figures PATENTEU FEB 2 5 ms SHEET 2 0F 2 532 $2 J; om fidz IN 334 OIX SlNnOO 1 PRODUCTION OF DISTILLERS YEAST CROSS-REFERENCE This application is a continuation-in-part of appli. cants copending US. patent application, Ser. No. 189,678, filed Oct. 15, I971 now abandoned.
BACKGROUND OF THE INVENTION 1. Field of the Invention This invention relates to a novel method for propagating yeast and, more particularly, to the propagation and storage of yeast used to inoculate distillery grain mashes in the production of whiskey and spirits from cereal grains and grain derivatives. By use of this method, a number of economic and operating advantages over the conventional methods of yeast propagation may be achieved.
2. Description of the Prior Art The conventional methods of propagating yeast for grain distillery use are fully described in numerous books and periodicals. One such reference is Industrial Fermentations, Chemical Publishing Co., New York, edited by L. A. Underkofler and R. J. Hickey. In Volume I, Chapter 2, by W. H. Stark, under Plant Yeast Procedure at page 45, the following statement is made: The memdium (for yeast propagation) is mash prepared at a higher concentration medium than the fermentor mash. Common practice is to use 16 gallons of-water per bushel of grain and (consequently) the yeast mash is relatively rich in nutrients as compared with a corn-spirits mash. At one time, either 100% barley malt mashes or 50% rye and 50% barley malt mashes were used. It has been found that a 70% corn to 30% barley malt mash provides-sufficient nutrients. Lower levels of malt may be inadvisable depending upon other. conditions. Thus, yeast mashes heretofore have contained appreciable amounts of barley malt (which is always more expensive than corn or milo) and rye (which is often more expensive than corn or milo).
Yeast mashes of ground rye and barley malt are generally heated in water slurry to gelatinize the starch and solubilize the other nutrients, such as the vitamins, minerals, and proteinaceous materials which are necessary for rapid yeast growth. After this mashing period, which permits these cereal enzymes to digest the grain starch and protein, the mash may either be immediately sterilized (sweet yeast mashing process) or the mash may be inoculated with lactic acid producing bacteria and incubated at l22130F. to produce 0.5-1 .5% lactic acid, then sterilized (sour yeast mashing process). After sterilization, the mash is cooled to the temperature desired for yeast growth, i.e., 7590F., and the yeast is introduced and allowed to grow until about 40-50% of the available sugars have been utilized. The yeast is then cooled to 50-60F. and can then be used for inoculating fermentors or it can be stored for a period of time needed but not longer than 24 hours as yeast cells die quite rapidly when propagated and stored as described above (see FIG. 1).
While the conventional methods for propagating yeast for grain distilleries may vary in certain details from that outlined above, the procedure is basically the same from distillery to distillery. Rye, barley, and barley malt are the usual grains used because, under the mashing conditions employed, these grains provide adequate amounts of the nutrients required to propagate distillers yeasts. Without special treatment, such as described herein, corn and grain sorghum (milo) do not support either rapid yeast growth or rapid growth and acid production by the lactic acid organisms used to sour yeast mashes. Therefore, mashes comprising high percentages of corn and/or milo, as normally pro cessed, cannot be used satisfactorily to propagate distillers yeast. Various additives to corn and milo mashes have been recommended to overcome these nutritional deficiencies, among them urea, inorganic nitrogen, protein concentrates, yeast cell preparations, vitamins, etc., but each of these adds to the cost of yeast propagation andoften has undesirable physiological effects upon the yeast or flavor effects upon the alcoholic distillate being produced.
The prior art also reveals other yeast growth supplements that have been proposed for cultivating distillers yeast. One example isfmold bran produced by cultivating certain species of Aspergillus on moistened, acidified bran (Underkofler, et al., Ind. & Eng. Chem. 38, 980-985, I946). These investigators recommend a yeast mash of about 94% com, 4% barley malt, and 1-2% mold bran along with inorganic nitrogen supplements, such as diammonium phosphate and ammonium sulfate, This method has the disadvantages of requiring special equipment for producing mold bran and, in addition, the use of at least 4% barley malt and nitrogen salts which add appreciably to the cost of yeast preparation.
Accordingly, it is a primary object of this invention to provide an improved method of propagating and maintaining distillers yeast.
An equally important object is to provide a method of propagating distillers yeast featuring rapid yeast growth, comparatively high viable cell counts, and exceptionally long yeast viability such that the yeast inoculum levels in alcoholic fermentation can be reduced by about one-half.
A further object is to provide a method of propagating distillers yeast on cereal grain substrates comprising at least about 95% com and/or grain sorghum (milo) and only minor amounts (i.e., 2% or less) of malted grain.
A still further object is to provide a method of propagating distillers yeastthat can be carried out with existing equipment and with existing facilities.
Yet another object is to provide an improved method of processing cereal grain to obtain whiskey and spirits.
SUMMARY OF THE INVENTION The foregoing and other objects, advantages, and features of the subject-invention may be attained by cultivating yeast for distillery grain fermentation on cereal grain mashes,- especially those comprising at least about 95%, preferably at least about 99%, corn or milo, subjected to action of glucamylase ferments such as those described in US. Pat. No. 3,418,211. It has been discovered that these ferments, which are rich in enzymes, especially amylases, proteinases, phytase, etc., have an exceptional capacity to modify cereal grain mashes for the growth and viability of distillers yeast.
More particularly, in accordance with the method of this invention, a cooked mash of ground cereal grain, especially corn and/or grain sorghum (milo) is prepared. The starch content of the cooked mash is thus gelatinized and, if desired, a minor amount (i.e., no
more than about 2% by weight) of salted cereal (e.g., barley malt) or microbial amylase may be added in order to thin and liquify the mash. Glucamylase ferment is added to the cooked cereal grain mash, acid producing bacteria or another source of acid may be added if desired, and the resulting mixture is incubated for about 4-6 hours and at a temperature of about l20-140F. in order to hydrolyze the starch and protein content of the mash. The treated mash is sterilized, cooled to about 80F., inoculated with distillers yeast and incubated at a temperature of about 80F. until the Balling drops about 4-5. The yeast culture is then cooled to about 60F. and stored in its propagation medium for up to3 days. The propagation of distillers yeast in accordance with the foregoing method 'has a number of advantages, principal among which are:
'l. more rapid yeast growth yielding larger numbers of active cells sooner;
2. more viability maintaining larger numbers of active cells longer;
3.1ess equipment, as inoculum size reduction permits more sets (i.e., inoculations) to be obtained from each propagation;
4. less raw materials, as fewer yeast tubs are required and rye and barley malt can be substantially eliminated;
5. less labor, as fewer yeast tubs need be prepared;
6. when propagated anaerobically, the yeast requires less time'to acclimatize to the final grain fermentation substrate;
7..more adaptability to automatic control because the substrates need not be specially prepared; and
. 8. operating steps have been greatly simplified.
IN THE DRAWING FIG. 1 is a plot yeast count versus time for the data DESCRIPTION OF THE PREFERRED EMBODIMENTS In accordance with this invention, the propagation and maintenance of distillers yeastcan be enhanced by subjecting the cereal grain mashes, on which the yeast is cultivated, to a glucamylase ferment which converts the nutrients of the cereal grain mashes to an available form that can be metabolized by the yeast.
More particularly, the method involves the steps of preparing a cooked mash of ground cereal grains, especially mashes comprising at least about 95% com and- /or grain sorghum (milo). The mash is cooked until the starch content is gelatinized, and a minor amount (i.e., no more than about 2%, preferably no more than about 1%) of malted cereal (e.g., barley malt) or microbial amylase may be added in order to liquify and thin the cooked mash. The resulting mash is then treated with glucamylase ferment and, if desired, with lactic acid producing bacteria, and the resulting mixture isfincubated for a time and at a temperature such that starch and protein of the mashare hydrolyzed (i.e., for about 4-6 hours at l20140F;) The mash is then sterilized,
cooled, inoculated with yeast, incubated and then cooled. The resulting incubated inoculum has higher than usual yeast-cell counts and greatly extended cell viability such that the cells maybe stored for 72 hours or longer before use.
tain 10-20% grain by weight, under submerged aerobic.
conditions with intensive aeration and agitation. The resulting ferment is called glucamylase ferment" because it heretofore has been primarily used in the rapid and near-complete saccharification of grain starch to glucose in alcoholic fermentations.
It has now been discovered that this type of ferment, which can be produced using several different species and strains of Aspergilli, has an exceptional capacity to modify cereal grain mashes for the growth and viability of distillers yeast. Among the Aspergilli suitable for the method of this invention are A. niger NRRL 330; A. foetidus NRRL 337; A. awamori NRRL 31 12 (also known as A. niger NRRL 3112); A. niger NRRL 3122; and A. oryzae NRRL 694, but the process of this invention is not limited to glucamylase ferments produced by these species or to the enzyme production methods described in US. Pat. No. 3,418,21 l. Forexample, enzyme products produced with selected cultures of Aspergilli by submerged aerobic cultivation such as those described by D. P. Langlois in US. Pat. No. 2,893,921, K. L. Smiley in US. Pat. No. 3,301,768, and H. E. Bode in [1.5. Pat. No. 3,249,514 could also be used to obtain glucamylase ferments for use in this invention. Likewise, enzyme products produced by cultivating selected strains of Aspergilli on moistened or semi-solid substrates or on the unagitated surface of liquid substrates could be used. Glucamylase ferments produced with A. awamori NRRL 3112 are preferred.
Glucamylase ferments are important for the subject process of yeast propagation because they make available yeast nutrients from grains, especially corn and/or milo,'thereby promoting yeast growth and viability. As a result, yeast cultures can be produced which possess exceptional numbers of active yeast and which maintain exceptional viability over long periods of time. Yet all of these features are attained more rapidly and at lower cost than by conventional methods.
Any of the several cereal grains such as wheat, corn, rye, barley, milo, and their derivatives may be used in the present method. Mixtures may also be employed. However, corn and/or milo are preferred by reason of their lower cost and by reason of the increased viability of yeast cells observed where corn and/or milo mashes are employed. Desirably, such mashes comprise at least about corn and/or milo. It is especially preferred to use at least about 99% corn and/or milo mashes, with less than 1% by weight barley malt or the equivalent in bacterial enzymes in order to liquify and thin the cooked ground cereal grain mash prior to treatment with glucamylase ferment.
The cooked cereal grain mash is prepared in a conventional manner. For example, a corn and/or milo mash containing about 18 gallonsof water per bushel of grain is cooked in water and/or stillage in order to gelatinize the starch. About 0.5% by weight barley malt is then added to liquify the mash. Alternatively, 200,000-400,000 D.V. units* of microbial amylase per *One D.V. unit dextrinizes 20 mg. Lintner Starch in 30 minutes at 30C. and pH 6.6. See Premier Malt Products, lnc., Bulletin dated Sept. 1, 1964.
The resulting mash, which now containsabout l bushel of grain per 25 gallons of volume is cooled to about 120l40F. and l,00020,000 units* (roughly about 0.5 to 1 pint) of glucamylase ferment are added per bushel of grain.
*A glucamylase unit is that amount of enzyme required to produce 1 gram of glucose from 4 grams of starch in 1 hour at 60C.
1f the preferred sour mashing process is desired, the mash is cooled to about l22-130F., preferably about 122F. prior to glucamylase addition, and about 2-10% by volume lactic acid culture (e.g., Lactobacillus delbrueckii) is also added. Enzymatic action and lactic acid production are allowed to proceed concurrently, generally for at least about 4 hours, preferably about 4-6 hours, at a temperature in the range of about l22-l25F., preferably about 122F. This is generally sufficient to lower the pH to about 5.0 or below and to liberate nutrients from the grain starch, protein, and other grain constituents.
If further souring is desired, the incubation at about 122."F. can be continued; if souring is adequate and further hydrolysis is desired, the temperature can be raised to at least about 140F.; if both are adequate and the mash is ready for inoculation with yeast, the temperature can be raised to around 160F., which terminates both souring and glucamylase action. The mash can be held aseptically at this temperature for a week or more if desired.
lnstead of using the lactic acid culture, an acidifying agent, such as stillage (the dealcoholized, liquid-grain residue from a previous grain alcohol fermentation) or sulfuric, hydrochloric, phosphoric, or lactic acid can be added prior to sterilization in order to reduce the mash pH to about 5.0 or lower as desired.
If a sweet yeast mashing process is desired, the cooked cereal grain mash can be treated with the glucamylase ferment in the range of about 120140F.
In any event, prior to inoculation with yeast, the mash treated as described above is sterilized at about 190F. or above for 2 hours or more, cooled to about 80F. (the yeast propagation temperature) and set with 2-3% yeast culture by volume. To obtain the high yeast counts that characterize this invention, the set Balling* should range between 18 and 21". As is known in the art, control of both the temperature and the Balling from this point on is .essential to making high viability yeast. As soon as the Balling has dropped 45 below set Balling, the mash must be cooled to 60F. or below and held at this temperature. This yeast culture is then ready to be used for inoculating grain mashes or it may be stored at this temperature for up to 3 days or more before using. Normally, the yeast culture is stored in its propagation medium, with portions being withdrawn Conveniently, the culture is maintained in the yeast tubs in which they are propagated.
*Balling is a measure of dissolved solids and is therefore a rough measure of sugar content.
As a result of the substantially longer viability of the yeast, it is ordinarily possible on a commercial basis to propagate only one or two batches of yeast per week, thereby greatly reducing yeast propagationcosts.
EXAMPLE 1 The improved growth and viability of distillers yeast which is obtained by the use of the present method over one of the older conventional methods have been demonstrated as follows. Hiram Walker Distillers Yeast No. 109 was inoculated into three different substrates, viz.: (l) 90% rye and 10% barley malt, which is commonly used in distillery practice, (2) 90% rye, 10% barley malt plus glucamylase ferment at the rate of 1 pint per bushel of grain, and (3) 99% com, 1% malt plus glucamylase ferment at the rate of about 1 pint per bushel of grain. These glucamylase ferments were produced with A. awamori NRRL 3112 in accordance with the method of US. Pat. No. 3,418,211. All mashes were soured with the same lactic acid culture and were then sterilized and cooled. Yeast culture was added and the mashes were incubated at the same temperature until the Balling was reduced 4. The mashes were then placed at 60F. and viable yeast num'berswere measured during the next 3 days by the usual method of serial dilution and plating in a medium containing 1% yeast Extract (Difco), 1% glucose and 1.5% agar adjusted to pH 6.8. Yeast colonies were counted after 48 hours incubation at 90F. The data are summarized in FIG. 1. It is apparent that the viable yeast count reached an appreciably higher figure in the 99% corn, 1% malt, glucamylase ferment mash than it did in either of the 90% rye, 10% malt mashes and these high viable counts persisted many hours longer. The glucamylase ferment improved the 90% rye, 10% malt mash-insofar as viable yeast counts were concerned but did not improve the yeast viability.
EXAMPLE 2 This experiment demonstrates that the subject method of anaerobically propagating distillers yeast in corn mash treated with glucamylase ferment produced with A. awamori NRRL 3112 results in a higher viable yeast population than does the treatment of corn mash from time to time for use in inoculating grain mashes.
with other enzymes, viz, barley malt or a commercial, purified amylolytic enzyme, i.e., Diazyme which is a product of Miles Laboratories, Inc., Elkhart, Ind. Diazyme, however, does possess some of the same type of stimulating activity.
Normal distillery corn mash, cooked on a 1 hour cycle reaching a maximum of 250F. and containing 1% barley malt for liquefaction of the mash, was divided into 3 portions of 500 ml each. The pH was adjusted to 5.0 with lactic acid and the mashes were sterilized at 250F. The mashes were cooled to F. and the following enzymic materials were added and allowed to act for 4 hours.
1. 4 ml (about 70 glucamylase units) of glucamylase ferment.
2. units of glucamylase as Diazyme.
3. 20 g. barley malt (about 2,000 Dextrinizing Units*). *A malt Dextrinizing Unit is that quantity of alpha amylase which will dextrinize soluble starch in the presence of an excess of beta amylase at the rate of 1 g/hr at 20C.
The mashes were then cooled to 60F., inoculated with by volume of actively growing yeast and incubated at 60F. for 18 hours. The viable yeast count was determined in each flask and the temperature was raised to 90F. After 5 hours, viable yeast counts again were made and the flasks were cooled to 60F. Thereafter, viable yeast counts were made at 24 hour intervals.
Results are shown in the table below:
VIABLE YEAST CELLS (millions/ml) (Flask l) (Flask 2) (Flask 3) Hours Glucamylase Diazyme Malt (cooled) 83 56 6l 24 at 60F. 162 120 129 48 at 60F. 270 250 160 72 at 60F. 370 2l0 150 96 at 60F. 320 I60 60 The viable yeast counts in the malt-treated mash are normal for distillery mashes. Treatment with the commercial enzyme, Diazyme, resulted in increased counts and improved viability, while treatment with glucamylase ferment resulted in both exceptionally high viable yeast counts and exceptionally long viability.
EXAMPLE 3 Flask No. Additive to 500 ml Corn Mash* l None 2 g. glucose 3 Vitamin mixture containing biotin. thiamine.
pantothenic acid and yridoxin 4 5 g. barley malt containing a total of 560 Dextrinizing Units of alpha amylase 5 0.5 g. papain (commercial grade) 6 40 units of glucamylase ferment (10 ml of A. awamori NRRL 3l l2 ferment) The glucose and vitamins were added after sterilization; the enzymes were added before the 5 hour incubation at 120F.
Distillers yeast inoculum was grown in corn mash I identical to flask l for 72 hours at 60F This culture was then used at the rate of 10 ml per flask to inoculate flasks 1 through 6. Viable yeast numbers were determined in each flask after inoculation and 5 hours incubation at 90F. Results were as follows:
It is apparent that the additionof vitamins and glucose had a low stimulatory effect on yeast growth in corn mash compared with the addition of these enzymes. Of the enzymes used, glucamylase ferment showed the highest stimulating effect on growth, although malt and papain showed some activity.
EXAMPLE 4 This experiment is an extension of Example-3 in that it compares the effects of enzymes. glucose. and vitamins on yeast growth and viability. Vitamins were included in this trial because they have long been known to increase the rate of growth of many microorganisms but have not been extensively studied for their effect on viability. The corn mash was prepared in a manner similar to Example 2 and was soured similarly with lactic acid organisms. The glucamylase ferment was prepared with A. awamori NRRL 3] 12. Below are the additives used:
Flask No. Additive to 500 ml Corn Mash Vitamins (same as under Example 3) l0 g. glucose (because this mash was semisolid due to its starch content, it was diluted with 1:] water) 5 g. barley malt containing a total of 560 Dextrinizing Units of amylase 0.5 g. papain (commercial grade) 40 units of glucamylase ferment 10 ml of ferment) All flasks were inoculated with distillers yeast and incubated overnight at 60F. The flasks were then placed at F. for 5 hours, then lowered to 60F. Viable yeast counts were determined as shown below:
Flask No. Additive to 500 ml Corn Mash Vitamins (same as under Example 3) 10 g. glucose (because this mash was semisolid due to its starch content. it was diluted with lzl water) 5 g. barley malt containing a total of 560 Dextrinizing Units of amylase 0.5 g. papain (commercial grade) 40 units of glucamylase ferment (10 ml of ferment) LII-I 0: [O
It is apparent from these results that glucose and vitamins have much less ability to promote the growth and maintain the viability of yeast in grain mashes than the enzyme additives. Of the enzymes used, glucamylase ferment was the most effective.
EXAMPLE 5 Previous experiments demonstrate the subject method of propagating yeast in corn mashes treated with certain enzymes. This experiment shows that yeast prepared by the subject method can be used at a lower than normal level to inoculate grain mashes to produce spirits or whiskey. A com spirits mash was prepared by cooking the corn with 0.5% barley malt (added for liquefaction) on a 1 hour cycle reaching a maximum of 250F. This mash was cooled to 148F. and 0.5% barley malt was added for further liquefaction and the mash was held at 148F. for 30 minutes. The mash was cooled to 90F. Stillage was added at the rate of 25% of the final volume. Glucamylase ferment. produced with A. awamori NRRL 31 I2. was added at the rate of 1 pint per bushel and distillers yeast grown on corn mash treated with glucamylase ferment was added at the rate of 0.5% of the final volume. Another fermentor of the same corn spirits mash prepared in the'same Hours of Storage Millions of Viable Yeast Cells/m1 manner was inoculated with 1% by volume of yeast i' g g gg$ grown on a yeast mash of 90% rye and 10%barley malt. These fermentors were followed for their rates of fer- 1; 38'- mentation and alcohol yields. Results are shown below: i; 48 366 70 27 270 Medium for Yeas B E It will be observed from these results that the corncmilucamylase glucamylase ferment mash supports much higher viable ermem at yeast numbers with much more extended viability than lnoculum 0.5 1.0 does the rye-barley malt yeast mash.
Viable yeast in inoculum when used millions/ml 115 100 EXAMPLE 7 This experiment illustrates that different species and Rate of fermentation,
weight loss in g. strains of Aspergllll produce ferments wh1ch are suit- 5g 2-8 gg able for the subject process. Ferments of A. awamori 43 hours NRRL 3112, A. niger NRRL 330, and A. foetidus 98 hours 3L8 NRRL 337 were prepared according to the process of Finalethylalcoholmm US. Pat. No. 3,418,211, and the resulting ferments tent, by weight 6.31 6.26 were used to propagate yeast as described under Examples 2 and 3 above, with these modifications: One thousand ml of corn mash was prepared for each using: 1
These results show that the yeast grown on cornglucamylase ferment, 3O glucamylase units of glucamylase ferment used at the rate of 0.5% by volferment produced Wlth A. awamo i NRRL 3112. ume fermented as rapidly and gave a yield of alcohol the Same amount fermfim used in 6x031)t that lhfi comparable to rye-malt yeast used at the rate of 1% by enzymes were inactiviated by heat before addition, (4) volume. about 80 glucamylase units produced with A. niger NRRL 330, and (5) about 65 glucamylase units pro- EXAMPLE 6 duced with A. foetidus NRRL 337.
This experiment illustrates the subject process under The'mashes were cooled to 60F. and inoculated with commercial operating conditions. A standard yeast 2% actively growing yeast by volume and incubated at mash of 90% rye and 10% barley malt was prepared by 60F. for 18 hours. Viable yeast counts were deter mashing these grains at 146F. for 30 minutes in a mined in all flasks and the temperature was raised to cooker, transferring to a yeast fermentor, cooling to 90F. After 5 hours, viable yeast counts again were adding la re and ng the mash at made and the flasks were cooled to 60F. Thereafter, l20-125F. The mash was then pasteurize cooled to viable counts were made at regular intervals. Results 80F., and inoculated with Hiram -Walker Distillers are shown below:
Number 1 2 3 4 5 Glucamylase None A. awumori A. awanzori A. niger A. foelidus Ferment (Active) (Inactive) Source Millions of Viable Yeast Cells/ml 0 Hours 5 7 5 5 7 5 Hours 90F.
(cooled) 43- 47 47 30 98 24 Hours 60F. 118 245 126 237 249 48 Hours 60F. 128 346 202 315 353 72 Hours 60F. 370 440 Yeast 109. After a- Balling drop of 4, the temperature was reduced to 60F.
A 99% com, 1% barley malt, glucamylase ferment yeast mash was prepared by cooking the corn and malt on a one hour cycle reaching a maximum of 250F., cooling to 122F., adding by volume 3% lactic culture and 1.1% glucamylase ferment (prepared with A. awamori NRRL 3112) and incubating until the mash reached the same pH (3.8) as the rye-malt mash described above. The mash was then pasteurized, cooled to 80F, inoculated with the same yeast strain and incubated until the Balling dropped 4 and then was cooled to 60F. Both mashes had similar grain concentrations before inoculating with yeast, i.e., about one bushel of grain in 25 gallons. Samples of each mash were removed periodically for the determination of viable yeast numbers with the results shown below:
It is apparent that ferments produced by different species and strains of Aspergilli are capable of stimulating yeast growth and improving yeast viability. The heat-inactivated ferment contained much less of this activity indicating that the active substances are enzymatic in nature.
glucamylase ferment. Rye-barley malt mashes inocu lated with this yeast strain and otherwise handled according to the previous examples give maximum viable cell counts of about 150 to 200 million per ml which decrease rapidly to about 50 million viable cells per ml in 72 hours.
EXAMPLE 9 In order to demonstrate that other distillers yeasts show similar growth and viability responses when cultivated by the subject process, the following experiment was performed with two additional distillers yeast strains, viz, Hiram Walker Distillers Yeast No. 5 and Hiram Walker Distillers Yeast No. 72. Two l-liter portions of corn mash were treated with glucamylase ferment and were simultaneously soured with lactic acid organisms. These mashes were pasteurized, cooled to 60F. and one mash was inoculated with 400 ml of anactively growing culture of Yeast'No. and the other with the same volume of an actively growing culture of Yeast No. 72. The inoculated mashes were held at 60F. overnight, then were raised to 90F. and incubation was continued at 90F. until a Balling drop of 4 to 5 was attained. These same yeast strains also were used to inoculate each of two l0-liter volumes of commercial rye-barley malt mash which had been cooked, soured, pasteurized, then incubated first at 70F. and then at 90F. until a similar Balling drop was attained.
- Thereafter, all four yeast mashes, two of each type,
were cooled to 60F. and held for 4 days. Viable yeast counts were determined at 24 hour intervals. Results are shown in the table below and in FIG. 2;
It will be seen from the data in the table and in FIG.
, 2 that these additional distillers yeast strains developed higher yeast cell counts with substantially longer viability when grown on corn mash treated withv glucamylase than when grown on rye-barley malt mash.
We claim: 7
l. A method of propagating distillers yeast comprising the steps of:
preparing a cooked mash of ground cereal grain in water with the cereal grain comprising at least 95% by weight of amember selected from the group consisting of corn, milo, and mixtures thereof;
adding to the mash glucamylase ferment produced by the submerged, aerobic fermentation of a cereal grain mash with a glucamylase producing mold strain selected from the group consisting of A. niger; A..f0etidus; A. awamori; and A. oryzae with about l,000-20,000 units of glucamylase ferment per bushel of grain being used to treat the cereal grain mash; V
incubating the resulting mixture of mash and glucamylase ferment at a temperature of about l20-l40F. for about 4-6 hours;
sterilizing the mash;
cooling the resulting sterilized mash to about F;
inoculating the resulting cooled sterilized mash with distillers yeast;
incubating the inoculated mash until the Balling of the mash has dropped about 45; cooling the resulting yeast culture to a temperature of no more than about 60F;
maintaining the yeast culture in the mash in which it was propagated at a temperature of no more than about 60F.; and storing theyeast at said temperature of no more than about 60F. in the mash in which it was propagated for at least about 72 hours and during storage withdrawing portions of yeast periodically to inoculate distillery grain mashes for alcohol production.
2. A method, as claimed in claim 1, wherein the cereal grain further comprises-an'enzymatic material selected from the group consisting of malted cereal grain and microbial amylase in an amount sufficient to liquify and thin the mash.
3. A method, as claimed in claim 1, wherein the pH of the mash is adjusted to a level no higher than about 5.0 prior to sterilization.
4. A method, as claimed'in claim.3, wherein the pH is adjusted by inoculating the mash with lactic acid producing bacteria, with the mash to which the glucamylase ferment and the bacteria have been added being incubated for at least about 4-6 hours at a temperature of about l22l25F.
5. A method, as claimed in claim 3, wherein the pH is adjusted by adding to the mash a member selected from the group consisting of stillage, sulfuric acid, hydrochloric acid, lactic acid, and phosphoric acid or mixtures thereof.
6. In the process for producing whiskey and spirits, the improvement comprising inoculating the cereal grain mash with about 0.52.0% by volume of a yeast culture produced in accordance with the method of